Bacillus anthracis, the cause of anthrax, has been developed into a weapon of mass destruction by foreign governments and terrorist groups. The recent release of B. anthracis in the United States, with severe consequences, exposed the need for a more effective response to this threat. B. anthracis is used as a weapon largely because it forms highly resistant spores that can be incorporated into explosive weapons and terrorist devices. Spores enter the body through skin abrasions and by ingestion or inhalation and then germinate and grow as vegetative cells. When growth occurs in internal tissues, the host usually dies within several days. Natural strains of B. anthracis are sensitive to common antibiotics; however, large-scale use of these antibiotics to protect against anthrax is logistically difficult and medically dangerous. In addition, antibioticresistant strains may be used in the future. Furthermore, the current vaccine for anthrax has proven problematic. Thus, new strategies are needed to respond to the anthrax threat, and these are likely to require detailed knowledge of the interactions between the mammalian immune system and the outermost surface of the B. anthracis spore - the exosporium. The exosporium serves as the primary interactive site with host defenses, as the source of surface antigens, and as a semi-permeable barrier that excludes antibodies and destructive enzymes. The exosporium consists of a paracrystalline basal layer and an external hair-like nap. Approximately 50% of the mass of the exosporium appears to be proteins, roughly 20 unique species including glycoproteins. Preliminary studies indicate that exosporium proteins play a role in spore virulence. The goal of this project is to determine the content, structure, and function of the external surface of the exosporium. Of primary importance will be the role in virulence of surface-exposed proteins and glycoproteins. Specifically, we will (1) identify these proteins, make mutations that alter them, and examine the effects of the mutations on host-cell interactions and virulence using a mouse model. (2) We will map epitopes on key exosporium proteins, such as the collagen-like protein (BclA) of the hair-like nap, and examine the effects of antibody binding to these epitopes. Of special interest are antibodies that bind exosporium proteins and inhibit spore germination. (3) We will determine the structure of Bc1A and its domains. (4) We will examine the structure and assembly of the basal layer of the exosporium, including the attachment of the hair-like nap. The entire program project is designed to provide sufficient understanding of spore-host interactions to enable the development of new treatments for anthrax.